Proton Transfer Pathways in Nitrogenase with and without Dissociated S2B

Abstract

Nitrogenase is the only enzyme that can convert N 2 to NH 3 . Crystallographic structures have indicated that one of the sulfide ligands of the active-site FeMo cluster, S2B, can be replaced by an inhibitor, like CO and OH À , and it has been suggested that it may be displaced also during the normal reaction. We have investigated possible proton transfer pathways within the FeMo cluster during the conversion of N 2 H 2 to two molecules of NH 3 , assuming that the protons enter the cluster at the S3B, S4B or S5A sulfide ions and are then transferred to the substrate. We use combined quantum mechanical and molecular mechanical (QM/MM) calculations with the TPSS and B3LYP functionals. The calculations indicate that the barriers for these reactions are reasonable if the S2B ligand remains bound to the cluster, but they become prohibitively high if S2B has dissociated. This suggests that it is unlikely that S2B reversibly dissociates during the normal reaction cycle.

Understanding the Electronic Structure Basis for N2 Binding to FeMoco: A Systematic Quantum Mechanics/Molecular Mechanics Investigation

“…Unfortunately, experimental characterization of dinitrogen-bound intermediates is scarce and the nature of the molecular and electronic structure of the cofactor prior to and after dinitrogen binding is unclear. Additionally, crystallographic studies − have shown that the bridging sulfide (S2B) between Fe2 and Fe6 can be displaced under certain conditions, implicating Fe2 and Fe6 as a site of ligand binding, but which could also be interpreted as sulfide lability being mechanistically relevant. , A recent study presenting an X-ray crystal structure apparently showing a missing sulfide and bound N 2 is controversial. − Some recent computational studies have investigated the protonation and dissociation of S2B and the mechanism of N 2 reduction after S2B dissociation, but it remains unclear whether S2B loss is part of the catalytic mechanism or not. − Other computational studies ,− have investigated N 2 binding to the whole cofactor; however, N 2 binding is rarely found to bind favorably to FeMoco. In a previous QM/MM study from our group, however, we found that E 4 -SP type cofactor models will favorably bind N 2 .…”

Biological nitrogen fixation in theory, practice, and reality: a perspective on the molybdenum nitrogenase system

“…It should be noted that an exo Fe-CH 3 group would likely be hydrolytically unstable and a detailed mechanism for regeneration of the E 0 resting state has yet to be clearly outlined [79]. Ryde has explored the N 2 ase reduction mechanism using computational studies, with recent work focusing on the potential role of the μ 2 -bridging S2B position in substrate binding and reduction [100][101][102]. In particular, a QM/MM approach (133 915 atoms) was paired with QM calculations (170-178 atoms), where two DF methods were employed: the non-empirical, pure functional TPSS and the hybrid B3LYP functional, to explore the optimized energies and geometries of a wide range of NNX-bound E 4 structures.…”

How Thermal Fluctuations Influence the Function of the FeMo-cofactor in Nitrogenase Enzymes

“…26 Using the BP86 GGA functional 27 , Hoffman and co-workers proposed an E 4 state with a "two-side" (2S) structure (Figure 1c), where the adsorption sites of hydrogen atoms occur on two sides of the bridged sulfurs [28][29][30] , to create a weakly bound N 2 28 Using a hybrid functional, Siegbahn has suggested a four-electron preactivation of the FeMo cluster, before the catalytic E 0 state is formed 31,32 (see also the supplemental information), but Hoffman et al have argued that such a proposed structure is inconsistent with experimental measurements 29 . Ryde's group carried out systematic calculations with different exchange-correlation functionals, particularly for the N 2 hydrogenation steps by considering the pathways with or without dissociation of the sulfur bridge [33][34][35][36] . Very recently, they validated the resting state (Fe II 3 Fe III 4 Mo III ) of E 0 with the calculation of redox potentials by quantum mechanical methods 37 .…”